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EL-MINIA MED., BULL., VOL. 19, NO. 2, JUNE, 2008
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HISTOLOGICAL AND IMMUNOHISTOCHEMICAL STUDY OF ACUTE
INCISIONAL WOUND HEALING IN MALE ALBINO RAT
By
Saadia Ragab, Azza Hussein, Seham Abd El-Raof and Hanaa Hassanein Mohammed
Department of Histology, Faculty of Medicine
ABSTRACT:
One of the most important functions of skin is tissue repair. This work was
carried to study the process of healing of acute incisional rat wound. Fifty five adult
male albino rats were used. Biopsies from 5 animals were used as control to study the
normal skin. The rest of the animals were wounded. Histological techniques include
hematoxylin and eosin stains and Masson trichrome stain for collagen.
Immunohistochemical techniques include the expression of cyclooxygenase II and
inducible nitric oxide synthase at different time points during wound healing.
The study of the wound with hematoxylin and eosin showed inflammatory cell
infiltration and creeping of new epidermal cells at 1 and 3 days postwounding. At 7,
14 and 30 days postwounding, a well established mature scar with thickening of the
newly formed epithelium above it was formed. Histological study of the wound with
Masson trichrome stain showed that a dense blue collagen network filled the wound
gap at day three postwounding. Mature distinct scar formation with compact
abnormally arranged collagen fibers was found at 7, 14 and 30 days postwounding.
Immunohistochemical study showed that high COX-2 expression at the wound site
started as early as 24 hours postwounding, continue to increase up to 7 days and
markedly decreased 14 days postwounding. High iNOS expression was found 24
hours and 3 days postwounding. Seven days postwounding iNOS expression
decreases and becomes faint at 14 and 30 days postwounding. It can be concluded that
wound healing starts by inflammation, reepithelialisation and ends by formation of a
scar of irregularly arranged collagen fibers. During the process of wound healing,
there was also the expression of inducible nitric oxide synthase (iNOS) and inducible
cyclooxygenase (COX-2). The two enzymes play important role in promotion of the
wound healing process.
KEY WORDS:
Skin
Healing
Wound
Histological
Collagen
COX2.
NOS
in inflammation, but also regulate other
critical
physiological
responses.
Currently three COX isoenzymes are
known, COX-1, COX-2 and COX-3.(Kis
et al., 2005).Cyclooxygenase-1 is
considered a constitutive enzyme, being
found in most mammalian cells. (Beiche
et al., 1996 and Amin et al., 1997). On
the other hand, Cyclooxygenase-2 is
undetectable in most normal tissues. It is
an inducible enzyme, becoming abundant in activated in cells at sites of inflammation (Chandrasekharan, 2002 and
INTRODUCTION:
Skin wound healing is a complex
process in which there is repair of
injuried epidermal and dermal tissues.
During wound healing, many inflammatory mediators were found (Theoret
2004, Desmouliere et al., 2005 and
Martin, 2005). Cyclooxygenase (COX)
is the key enzyme required for the
conversion of arachidonic acid to prostaglandins (PGs) which are compounds
that present in a wide variety of human
tissues. PGs not only play a central role
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Warner et al., 2002). Nitric oxide is a
very important signaling molecule that
acts in many tissues to regulate a wide
range of physiological processes. (Witte,
2002). It was first discovered several
years ago by a group that was attempting
to identify the agent responsible for
promoting blood vessel relaxation and
the regulation of vascular tone. This
particular agent was named endothelium-derived relaxing factor (EDRF),
and was initially assumed to be a protein
like most of the other signaling factors
previously discovered. The discovery
that EDRF was in fact nitric oxide, a
small gaseous molecule that plays a key
role in many biological proc-esses
(Gryglewski, 1986). Nitric Oxide (NO)
is produced by a group of enzymes
called nitric oxide synthases (NOS).
There are three isoforms of NOS which
have been named according to their
activity. They are neuronal NOS
(nNOS), endothelial NOS (iNOS), and
inducible NOS (iNOS) (Palmer, 1988).
The aim of this study is to investigate the
morphological changes in cells and
tissues as a result of acute incisional skin
wound and to evaluate the expression of
some inflammatory mediators during
acute incisional wound healing in male
albino rat using histological and immunohistochemical techniques.
were used as control to study the
normal skin. The rest of the animals
were wounded, briefly, the dorsal skin
was shaved and swabbed with 70%
alcohol, then one 1 cm full thickness
incision was made on the back of each
animal according to a template (Fig. 1,
A). The wounds were unsutured and
allowed to heal by secondary intention.
The animals were housed in individual
cages and allowed to recover. Animals
were killed at 1, 3, 7, 14 and 30 days
postwounding (10 at each time point)
by chloroform overdose followed by
cervical dislocation and animals were
shaved if necessary. The cephalic and
the lumber regions of the wound site
were marked using non toxic marking
pin and photographs of the wounds
were taken (Fig. 1, B). The dorsal skin
was then removed from the animals by
using a sterile surgical blade down to
and including the panniculus carnosus
muscle; from each rat, one wound was
dissected out with half a cm from the
surrounding non-wounded surrounding
skin and then bisected perpendicular to
the plane of the wound, one half was
used for histology and the other half
was used for immunostaining. The
specimens were fixed using 10%
formal saline for 48 hours. After
proper fixation, the specimens were
dehydrated, cleared, impregnated with
soft paraffin and then embeddd in hard
paraffin and 6 microns sections were
cut using rotatory microtome for
histological staining. and the following
methods were used:
MATERIAL AND METHODS:
This study was based on
biopsies taken from the skin of 55
adult male Sprague Dawley rats. The
animals aged approximately 12 weeks
old and weight 225 to 250 gm. They
were matched and singly housed in
plastic cages and maintained in a light,
humidity and temperature controlled
environment for one week prior to the
experiment. Standard rat diet and water
were allowed. After wounding, animals
were kept under the same conditions
until the end of the experiment.
Animals were briefly exposed to
chloroform. Biopsies from 5 animals
Histological techniques: Hematoxylin
and eosin: stain and Massons
trichrome stain.
Immunohistochemical techniques:
Cyclooxygenase-2 (COX2) antibody
and inducible nitric oxide synthase
(iNOS) antibody.
Prior to immunolabelling, sections were fixed in acetone for 10
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minutes. Endogenous peroxidases
were quenched by treatment with 0.5%
H2O2 in methanol followed by
washing in tris buffer saline(TBS).
Non specific binding of IgG was
blocked using normal goat serum,
diluted in 1:50 in 0.1% bovine serum
albumin in TBS for 30 minutes. The
sections were incubated with the
diluted primary antibodies (iNOS 1:
1000 and COX2 1:500) at 4C overnight. Sections were then washed 3
times 5 minutes each and incubated for
another 30 minutes with biotinylated
secondary antibodies diluted 1:1000,
followed by washing. Following a
further 30 minutes incubation with
Vectastin ABC kits (Avidin, biotinylated horse radish peroxidase complex)
and washing for 10 minutes, the substrate, diaminobenzidine tetrahydrochloride (DAB) in distalled water was
added for 5-10 minutes. The slides
were dehydrated and mounted. This
results in blue staining in positive
sites.
could be noticed at day three
postwounding and the creeping new
epidermal cells were also observed (Fig.
3). At 7 days postwounding, there was
full reepithelialization with increase in
the newly formed epithelial thickness
and narrowing of the wound with matrix
condensation leading to closure of the
gap created by the injury and scar
formation. Lack of hair follicles and so
other skin appendages in the scar was
also observed. More maturation in granulation tissue and decrease in the inflammatory cell infiltration were basic
findings at this time point (Fig. 4).
At 14 and 30 days wounds
show a well established mature scar
with thickening of the newly formed
epithelium above it by comparison to
the surrounding normal skin. The
absence of hair follicles and other skin
appendages at the scar site are still
present at these time points (Figs. 5 &
6). New blood vessels beneath the scar
were seen at the age of 30 days
postwounding (Fig. 6).
RESULTS:
Haematoxylin and eosin:
The epidermis of thin skin
showed a single stratum germinativum
as the thick skin however it contained a
thin stratum spinosum and stratum
corneum. Thin skin lacked a definite
stratum lucidum and stratum granulosum, although individual cells of these
layers were present in their proper
locations. Thin skin contained a variety
of appendages, mainly hair follicles,
sweat glands, and sebaceous glands
(Fig 1).
Massons trichrome:
Collagenous connective tissue
of the dermis appears blue, parallel to
the surface of the skin and arranged in
honeycomb architecture. Elastic fibers
appeared also in the dermis and stained
red with Masson trichrome. They
appeared especially abundant near
sebaceous and sweat glands. Groups of
smooth muscles were located in the
deeper regions of the dermis and
stained red also with Masson trichrome
(Fig. 7). Wounds at early time points
one day postwounding were covered
by blood clot and the cut was through
the whole thickness of the skin (Fig.
8).At day three postwounding, a dense
collagen network stained blue by
Masson trichrome filled the wound
gap. Wounds showed abnormal collagen orientation at the wound site as
compared to the normal orientation in
At the earliest time point at day
one postwounding there was blood clot
over the wound and a provisional matrix
with inflammatory cell infilt-ration. The
start of creeping of new epidermal cells
could be observed (Fig. 2). Marked
increase in the inflammatory cell infiltration with degradation of the blood clot
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the normal skin surrounding the
wound. At 7 days postwounding: the
fibrin-rich matrix was replaced by
dense collagen staining. It showed
condensation of the matrix and scar
tissue formation which started to
contract leading to narrowing of the
wound (Fig. 9). Scars at 14 and 30
days postwounding showed increase in
matrix condensation and mature distinct scar formation with compact
abnormally arranged collagen fibers.
At this stage the wound had remodelled to form a scar (Figs. 10 & 11).
infiltrating the wound (Fig. 15). At 7
days postwounding, there was marked
reduction in COX2 expression. It was
found in the few inflammatory cells
near the scar and in the endothelial
cells of the blood vessels (Figs. 16 &
17). Scars at 14 and 30 days postwounding show faint COX2 expression. It could be observed in the
thickened epithelium above the scar
and in the endothelial cells of blood
vessels deep in the scar. It was hardly
seen in the scar and even less than in
the surrounding normal skin where it
was expressed mainly in hair follicles
(Figs. 18 & 19). Wounds at early time
points one day postwounding showed
high iNOS expression at the wound
site mainly localized to the creeping
new epidermal cells at the wound
edges and the inflammatory cells infiltrating the wound (Fig. 21). Marked
increase in iNOS expression was
observed three days postwounding
localized mainly to the creeping new
epidermal cells at the wound edges
and the inflammatory cells infiltrating
the wound (Fig. 22). There was little
expression of iNOS in the keratinocytes as compared to the high expression in the inflammatory cells (Fig.
22,A). Expression of iNOS was also
seen in the endothelial cells lining the
blood vessels and the perivascular mast
cells (Fig 22 B&C). At 7 days postwounding, there was marked reduction
in iNOS expression. It was observed in
the few inflammatory cells near the
scar and in the endothelial cells of the
blood vessels (Fig. 23).Scars at 14 and
30 days postwounding showed faint
iNOS expression. It could be seen in
the thickened epithelium above the
scar. It was hardly seen in the scar and
even less than in the surrounding
normal skin where it was expressed
mainly in hair follicles (Figs. 24 & 25).
Immunocytochemistry:
Cyclooxygenase-2
(COX2)
expression appeared brown in positive
staining. Weak expression of COX2
was found in the normal rat skin. It
was mainly observed in hair follicles
and in the epidermal cells (Fig. 12),
inducible nitric oxide synthtase (iNOS)
expression appeared brown in positive
staining. Weak expression of iNOS is
seen in the normal rat skin. It was
mainly localized in hair follicles and in
the epidermal cells (Fig. 20). Wounds
at early time points one day
postwounding showed high COX2
expression at the wound site mainly
localized to the creeping new
epidermal cells at the wound edges
and the inflammatory cells infiltrating
the wound (Fig. 13). There was little
expression in the keratinocytes as
compared to the high expression in the
inflammatory cells (Fig. 14,A). COX2
expression was also observed in the
endothelial cells of blood vessels (Fig.
14, B). The high COX2 expression in
mast cells and fibroblasts was also
noticed at this time point (Fig. 14,C).
Marked increase in COX2 expression
was noticed three days postwounding.
It was mainly localized to the creeping
new epidermal cells at the wound
edges and the inflammatory cells
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Figure 1. photomicrograph showing normal thin
skin with thin epidermal layers ( arrow), hair
follicles (double arrows) with associated sebaceous
glands (arrow with round head) and thin layer of
keratin . (H&E) x40
Figure 2. photomicrograph of rat dermal wound
1day postwounding. showing the blood clot over the
wound (star),
inflammatory cells infiltrating the
wound ( arrow) and the creeping new epidermal
cells ( double arrows). (H&E x40.
Figure
3.
photomicrograph
of
skin
3
days
postwounding showing the marked increase in the
inflammatory cell infiltration at the wound site
(arrow), the creeping new epidermal cells ( star) and
the degradation of the blood clot .(H&E )x40.
Figure
4.
photomicrograph
of
skin
7
days
postwounding showing closure of the wound,
reepithelialisation with thicknening of the epithelium
(arrow), condensation of the matrix and the scar
tissue formation ( S). H&E x10
Figure 5. photomicrograph of skin 14 days
postwounding showing the scar (S) with the thick
epithelium over it (arrow). (H&E) x10.
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Figure 6. photomicrograph of skin 30 days
postwounding. A) showing the scar (S), the loss
of hair follicles , neovascularization (arrow) and
absence of skin appendages in the H&E x40
Figure 7. photomicrograph of normal rat skin
showing blue collagen fibres organised in a
‘honeycomb' formation in the normal dermis (
arrows) and the red colour of the muscle (double
arrows). Masson trichrome x10
Figure 8. photomicrograph of 1 day dermal
wound showing the junction between the full
thickness wound with the blood clot over it
(black arrow) and the normal skin (blue
arrow). Masson trichrome x10
Figure 9. photomicrograph of 7 days dermal
wound showing condensation of the matrix and
scar formation (s). Masson trichrome x10.
Figure 10. photomicrograph of 14 days dermal
wound showing connective tissue fibres mainly
collagen are densely packed to form a mature scar
(S). The collagen contrasted sharply with the
adjacent nomal dermal collagen. Masson trichrome
x10.
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Figure 11. photomicrograph of 30 days dermal
wound showing connective tissue fibres mainly
collagen are densely packed to form a mature
scar . Masson trichrome x10.
Figure 12. Immunohistochemical evidence of
COX2 expression in normal rat skin. There is faint
COX2 staining mainly in the epidermal cells
(arrow) and hair follicles (double arrows).x10.
Figure 13. Immunohistochemical evidence of
COX2
expression
1
day
postwounding
showing high COX2 expression at the wound
site mainly localized to the creeping new
epidermal cells at the wound edge ( arrow)
and the inflammatory cells infiltrating the
wound (double arrows). x10.
Figure 14. Immunohistochemical evidence of COX2 expression 1 day postwounding. A) Showing
COX2 expression in the skin at the wound edge. Notice the little expression in the keratinocyte
(K) and the high expression in the inflammtory cells (arrow) in the dermis. B) COX2 is seen in
the endothelial cells of blood vessels (arrow). C) COX2 is seen in mast cells (red arrows) and
fibroblast like cells (black arrows) at the wound site. x40.
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Figure15.
Immunohistochemical
evidence
of
COX2 expression 3 day postwounding. High
expression of COX2 in the growing newly formed
epithelium at the edge of the wound ( arrow) and
in the inflammatory cells infiltrating the wound
(double arrows). x10.
Figure 16. immunocytochemical evidence of COX2
expression 7 days postwounding showing marked
reduction in COX2 immunoreactivity at the scar
site (s). Few inflammatory cells expressing COX2
at the edges of the scar (arrows). x10.
Figure 17. immunocytochemical evidence of COX2
expression 7 days postwounding showing COX2
expression in the endothelial cells of blood vessels
(arrows). x10.
Figure 18. Immunohistochemical evidence of
COX2 expression in
Faint
COX2
14 days postwounding.
expression
in
the
thickened
epithelium above the scar (arrow) and deep in
the dermis (double arrows). x10.
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Figure 19. Immunohistochemical evidence of COX2
expression 30 days postwounding. COX2 is expressed
in the endothelial cells of vessels deep in the scar (
arrow). It is hardly seen in the scar and even less than
in the surrounding normal skin where it is expressed
mainly in hair follicles (double arrows). x10.
Figure 20. Immunohistochemical evidence of iNOS
expression in normal rat skin. The ABC technique
was used and it shows brown in positive staining.
There is faint iNOS staining mainly in the epidermal
cells ( arrow) and in the hair follicles (double
arrows). x10.
Figure 21. Immunohistochemical evidence of iNOS
expression 1 day postwounding showing high iNOS
expression at the wound site mainly localized to the
growing epithelium at the wound edge (arrow) and
the inflammatory cells infiltrating the wound
(double arrows). x10.
Figure 22. Immunohistochemical staining showing cellular localization of iNOS in rat dermal
wound healing. A) iNOS is faintly expressed in keratinocytes (K) of the epidermis next to wound
area, in inflammatory cells (arrow) infiltrating the dermis close to the wound edge. B) in the
vascular endothelial lining (long arrow) and the perivascualr mast cells (short arrow) at the
wound site. C) in the vascular endothelial cells (arrows). A x10, B and C x40.
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Figure 23.
Immunohistochemical evidence of iNOS
expression 7 days postwounding showing marked
reduction in iNOS immunoreactivity at the scar site
(S). Few inflammatory cells expressing iNOS at the
edges of the scar (black arrows). iNOS is also
expressed in the endothelial cells of the blood vessels
Figure 24.
Immunohistochemical evidence of iNOS
expression
14
days
postwounding.
Faint
iNOS
expression in the thickened epithelium above the scar
(black arrow) and deep in the dermis (red arrows).
x10.
Figure 25. Immunohistochemical evidence of iNOS 30
days postwounding. It is hardly seen in the scar (s) and
even less than in the surrounding normal skin where it
is expressed mainly in hair follicles ( arrow). x10.
in the wound area included inflamematory cell infiltration, blood clot
formation, beginning of reepithelialisation and loss of the normal collagen
orientation. These changes were in
agreement with those of (Flanga,
2005). This was in favourite of the
healing process since blood clotting
stops excessive bleeding (Romo and
Pearson, 2005). Reepithe-lialisation
aimed at closing of the wound to
enhance the healing process (Garg,
2000). The changes in the wound area
DISCUSSION:
This study aimed at the investigation of the histological changes that
occur in rat skin during healing of
acute incisional wounds at different
time points 1, 3, 7, 14 and 30 days
postwounding. The work investigated
also the expression of inflammatory
mediators cyclooxygenase (COX-2)
and inducible nitric oxide synthase
(iNOS), which are known to contribute
to the pathogenesis of wound healing
and scar formation. The changes found
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continued to occur throughout the time
course; three days postwounding the
wound showed degradation of the
blood clot and marked increase in the
inflammatory cell infiltrate, this was in
agreement with (Shah et al., 1999). By
day 7 postwounding the healing process was almost completed with complete reepithelialisation, scar formation
and retraction of the cut muscle edges
leading to narrowing of the wound. By
day 14 the scar was maturated. These
findings were in accordance with the
results of other investigators ((Foreman
et al., 1996). Although reepithelialisation started as early as one day postwounding, it increased with time and
the wound showed complete reepithelialisation seven days postwounding,
this was in agreement with findings of
(Shah et al., 1999). Many studies
(Ferguson et al., 2001 and Santoro,
2005) including the present one have
shown that although the course of
wound healing was controlled by three
main phases inflammation, reepithelialisation and collagen deposition,
there was overlapping between these
three
phases
and
there
was
predominance of each phase at certain
time point; inflammation predominates
in the first three days postwounding.
Reepithelialisation predominates seven
days postwounding. Collagen deposition predominates late in wound
healing and this coincides with the
results of (Sirsjo, 1996). It starts as
early as 3 days postwounding and
continued over the time course till it is
fully mature by 30 days. The white
distinct appearance of the scar is due to
the loss of the normal collagen
orientation in the wound area
(Chamberlain et al., 1994). The
induction of iNOS and COX-2 at the
wound was during the early time points
1-7 days postwounding. Therefore, it
was coincident with the infiltration of
the wound by inflammatory cells and
induction of cytokines. There was
earlier induction and earlier downergulation of iNOS in the wounds by
comparison to COX-2. While iNOS
was highly expressed mainly during
the first 2 days postwounding, COX-2
was highly expressed 2-7 days wound.
This could be explained by (McCall &
Vallance, 1992) who mentioned that
iNOS is expressed by neutrophils, the
first effector cells which migrate into
the wound and predominate during the
first 24 hours after injury. Following
appropriate stimuli, neutrophils can
express cytokines including interleukin
-3, IL-1, IL-12, IL-8 and tumor
necrosis factor (TNF), which are
known to induce iNOS. However,
COX-2 were expressed by fibroblasts
and mast cells, which infiltrate wounds
at late time point (sirsjo,1996). In
addition, (Rosenberg, 2006) documented that transforming growth factor
(TGF) and cytokines which is known
to be upregulated in wounds at late
time point can inhibit iNOS and
induce COX-2. Other investigators
(Salvemini et al., 1989) added that
iNOS stimu-lates COX-2 and this
could explain the lag in COX-2
activity. In summary, there are several
factors, which could control the expression and the profile of iNOS and
COX-2 in the wounds, including the
wound cellularity, growth factors and
the products of the enzymes.The
pattern of expression of iNOS in the
wound could be a physiological
requirement for healing. Nitric oxide
has anti-microbial and anti-proliferative activities which could overcome
the microbial contamination in the
early stages after injury (Nussler &
Billiar, 1993). Moreover, nitric oxide
produced by the vascular endothelial
cells, maintains a high rate of blood
flow in the wound. This increased blood
flow facilitates both nutrition and cell
infiltration. Further-more, this low level
of nitric oxide could neutralize the
oxygen free radicals released by
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infiltrating leukocytes, thus protecting
cells from damage by oxygen and
hydroxyl radicals (Romo and Pearson,
2005). On the other hand, cytokines
released by infiltrating macrophages
and associated with bacterial invasion
several hours postwounding, could
stimulate poly-morphnuclear neutronphils (PMNs) and macrophages to
produce large amounts of nitric oxide,
which in turn might inhibit cytokine and
nitric oxide produ-ction by infiltrated
cells (Griscavage et al., 1993). This
could provide a negative regulation of
the inflammatory response of the
wound, since severe inflame-mation
and high levels of nitric oxide can cause
massive tissue damage. Transforming
growth factor  (TGF), which is
known to be a potent fibro-genic
cytokine that enhances wound healing,
has been reported to inhibit nitric oxide
production by macrophages (Eichler
and Carlson, 2005).The PGs are known
to have anti-inflammatory effects and
Recently, (Kis et al., 2005) found that
PGs enhance wound repair by
enhancing blood flow in the neovascularisation of repairing wounds.
Thus the course of wound healing is
normally controlled by both iNOS and
COX-2 and it is likely that, interference
with these physiological responses to
injury either by inhibitors or inducers
for these enzymes could alter the
healing process. However delayed
healing which occurs under conditions
associated with low expression of the
enzymes e.g. in diabetes and
malnutrition (Schaffer et al., 1997) or
excessive upregulation of these enzymes such as chronic wounds might be
treated using inhibitors or inducers
(Flanga, 2004 and Scholar,2006). (Koh
et al., 2003) showed that COX-2
activity was increased during early
time points of wound healing. Using
immunostaing they showed that COX2 staining was particularly prominent
in the inflammatory cells, fibroblast
and newly formed vascular endothelial
cells. These data suggest that COX-2 is
constitutively expressed in epidermis
and is associated with keratinocyte
differentiation.
Pervious
studies
(Etscheid et al., 2005)) observed that
the expression of COX2 was mainly
within the basal layer of the epidermis,
peripheral cells in the outer root sheath
of hair follicles and fibroblast-like cells
and capillaries near epidermal wound
edges. Much less intense expression
was observed in normal skin than in
injured skin. (Ferguson et al.,2001)
added that administration of the COX2 inhibitor delayed re-epithelialization
in the early phase of wound healing.
Thus, COX-2 induction may be
important in cutaneous wound healing
through enhancing reepithelailization.
From all of the above, it can be concluded that wound healing is a complex
process that includes inflammation,
reepithelialisation and ends by formation of a scar of irregularly arranged collagen fibers. During the process
of wound healing, there was also the
expression of inducible nitric oxide
synthase (iNOS) and inducible cyclooxygenase (COX-2). The two enzymes
pay an important role in promotion of
the wound healing process.
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‫الدراسه الهستولوجيه والمناعه الكيميائيه الخلويه‬
‫اللتئام الجرح الحاد القاطع لذكر الفأراالبيض‬
‫سعديه رجب‪ -‬عزه حسين على‪ -‬سهام عبد الرءوف – هناء حسانين محمد‬
‫قسم الهستولوجى ‪ -‬كلية طب المنيا‬
‫ان أحد اهم وظائف الجلد اصالح النسيج‪ .‬وقد تم هذا العمل لدراسة عملية التئاام الجارح‬
‫الحاد القاطع للفئران وقد استخدم لذلك خمسة وخمسون من ذكورالفئران البيضاء البالغة‪.‬وتم اخد‬
‫عيناا مان خمساة ائاران كمجموعاة ضاابطة وتام قتال بقياة الفئاران بعاد جرحاااا اليااوم ا ول‬
‫والثالث والسابع والرابع عشار والثالثاين ‪.‬وقاد اوضاح دراساة الجارح بصابغة الايماتوكسايلين‬
‫وا يوسين ان نشع خاليا ا لتاا وححف خاليا البشارا الجديادا يكاون اا الاثالث اياام ا ولا‬
‫بعد الجرح‪ .‬وا ا يام السابع والرابع عشر والثالثين تم تكوين ندبا ناضاجا حسان التاسايع ماع‬
‫حياادا ساامك النساايج الطالئااا حااديث التكااوين اوقااه‪ .‬امااا الدراسااه الاسااتولوجيه للجاارح باسااتخدام‬
‫صبغة الماسون تراى كروم اقد بينا ان شابكه كثيفاه مان الكاو جين المصابوزة باا حر تماال‬
‫اجاوا الجاارح عناد اليااوم الثالا ث‪ .‬اماا اااا ا ياام السااابع والرابااع عشار والثالثااين اقاد وجااد النااد‬
‫الناضج المكون من الياف الكو جين الملتحمه المترتباه زيار طبيعياا‪ .‬وقاد بينا دراساة المناعاة‬
‫الكيميائية الخلوية ان التعبير المرتفع نحيم السيكلواوكسجينيح اثنين عند مكان الجرح يبدا مبكرا‬
‫بعد اربع وعشرين ساعة من الجرح ويستمر ا ا رتفاع حتاا سابعة اياام ويقال جادا عناد الياوم‬
‫الرابع عشر والثالثينن ويكاون هاذا ا رتفااع علاا ا خاك ااا كال مان النسايج الطالئاا حاديث‬
‫التكوين عند حااتا الجرح‪ ،‬والنسيج المتحب اا وسط الجرح ‪،‬وا وعية الدموية حديثة التكوين‬
‫وخاليا ا لتاا التا تنش ع اا الجرح‪ .‬ويكون التعبير قليل للغاية ااا البشارا وا دماة بعيادا عان‬
‫مكان الجرح‪.‬‬
‫وقااد كااان التعبياار مرتفعااا نااحيم النيتريااك اوكساايد سااينثيح المسااتحث اااا اليااومين ا ول‬
‫والثالااث بعااد الجاارح ووجااد علااا ا خااك اا كاال ماان الخاليااا المبطنااه لالوعيااة الدمويااه حديثااة‬
‫التكااوين والنساايج الطال ئااا‪ .‬ولكنااه يقاال جاادا اااا نساايج النااد عنااد ا يااام السااابع والرابااع عشاار‬
‫والثالثين‪ .‬ويمكن من ذلك استنتاج ان عملية التئام الجروح تشتمل علا ا لتااا واعاادا النسايج‬
‫الطالئا وتكوين الند من اليااف الكاو جين المترتباه ااا صاور زيار طبيعياه‪ .‬وتشاتمل ايضاا‬
‫علا تعبير كل من انحيم السيكلواوكسجينح اثنين وانحيم النيتريك اوكسيد سينثيح المستحث حيث‬
‫ان لاما دورا ماما ا تحفيح عملية ا لتئام‪.‬‬
‫‪15‬‬